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Abstract In this study, the gene encoding for bovine interferon-tau (bIFN-τ), with signal sequence, was obtained through PCR from bovine early embryos and subcloned into pGEM-T vector. After being verified, the bIFN-τ fragments with signal sequence or without signal sequence were inserted into the expression vector pET-30a(+), respectively. Two recombinant plasmids were induced to express the recombinant proteins by Isopropyl β-D-1-thiogalactopyranoside, respectively. The expressed products were then denatured, renatured and purified. The biological activities of purified rbIFN-τ were identified by assaying its antiviral activity and its effect on the morphology of bovine endometrial epithelial cells during in vitro culture. The results showed that bIFN-τ gene could be obtained from five bovine blastocysts by PCR without extraction of genomic DNA. Its homologies were 99% in nucleotide acids and 97% in amino acids to the sequence in GenBank (XM593584). The products of rbIFN-τ with minus signal sequence expressed in pET-30a(+) were analyzed by SDS-PAGE, and a new protein of 20kD was detected. Its molecular weight was as the same as the designed. The antiviral activity of rbIFN-τ was 1×104IU/mg using standard cytopathic reduction assay. The rbIFN-τ induced obvious morphological changes in
bovine endometrial epithelial cells in vitro. The cell volume was larger than the control and a lot of vesicles appeared in the cytoplasms after 24 h culture in presence of 2.9µg/ml rbIFN-τ. In conclusion, the purified rbIFN-τ with biological activities was obtained in this experiment.
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Received: 11 July 2007
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